The present invention relates to compounds which bind to all or parts of the active binding xe2x80x9csouth pole pocketxe2x80x9d of the LFA-1 I-domain and their uses as LFA-1 antagonists.
The lymphocyte function associated antigen LFA-1 belongs to the xcex22-integrins and plays an important role in T-cell activation and extravasation. Interactions of LFA-1 with its counter-receptors on endothelial and antigen presenting cells such as ICAM-1 or ICAM-3 are an important process in leucocyte endothelial cellular adhesion and migration which mediates disorders or diseases, e.g. autoinmuune diseases, inflammation, ischemia/reperfusion injury and graft rejection after transplantation.
The so-called I-domain (Inserted Domain) of LFA-1 comprises a module of about 190 amino acids (Takada et al., Matrix Biology, 16, 143-151, 1997). The I-domain folds into a common structural motif comprising a central xcex2-sheet surrounded by helices as determined by X-ray crystallography. (A. Qu and D. Leahy, Proc. NatI. Acad. Sci. USA, 92, 10277-10281, 1995).
Compactin and Mevinolin are fungal metabolites which have following formula: 
They are disclosed e.g. by Y. Chapleur in xe2x80x9cProgress in the Chemical Synthesis of Antibiotics and Related Microbial Productsxe2x80x9d, Springer Verlag, 1993, vol. 2, 829-937.
Mevinolin and most of the known analogues, e.g. pravastatin, mevastatin, simvastatin etc. have been found to be useful e.g. as 3-hydroxy-3-methyl glutaryl coenzyme A reductase (HMG CoA R) inhibitors.
In accordance with the present invention, it has now surprisingly been found that mevinolin and derivatives thereof bind to LFA-1. Accordingly, the invention provides compounds for use in the treatment or prevention of autoimmune diseases, inflammation, ischemia/reperfusion injury and graft rejection which are preferably specific or substantially specific LFA-1 binding molecules, e.g. specific or substantially specific inhibitors of LFA-1/ICAM-1 or ICAM-3 interactions. Such compounds are preferably other than LFA-1 antibodies.
More particularly, it has been found that mevinolin binds to the LFA-1 I-domain between the C-terminal helix xcex17 and one side of the xcex2-sheet (hereinafter the xe2x80x9csouth pole pocketxe2x80x9d). X-ray analysis of the complex of LFA-1 I-domain with mevinolin shows that mevinolin does not bind to the MIDAS-site (xe2x80x9cmetal ion dependent adhesion sitexe2x80x9d).
The complex, LFA-1 I-domain/mevinolin, is prepared by adding mevinolin (100 mM solution in DMSO) to the protein solution (12.7 mg/ml, 100 mM MgSO4), followed by crystallization. The structure is solved by molecular replacement (using the coordinates of apo LFA-1 I domain, A. Qu and D. Leahy, above) and has been refined to a R factor of 19.4% (Rfree=25.9%) using X-ray amplitudes in the resolution range 8 xc3x85-2.6 xc3x85. The final model contains 2xc3x97182 amino acids (amino acid residues 128 to 309 of the xcex1-chain of LFA-1 which corresponds to the I-domain), 2 mevinolin molecules and a total of 86 water molecules.
The south pole pocket is a cavity between one side of the central xcex2-sheet (amino acids of xcex21, xcex23, xcex24, xcex25, preferably the side chains of such amino acids) and the xcex1-helices xcex11, xcex17 (secondary structure of LFA-1 I-domain). Preferably the south pole pocket is the cavity defined by amino acids Val 130, Leu 132, Phe 134, Phe 153, Val 157, Leu 161, Tyr 166, Thr 231, Val 233, Ile 235, Ile 255, Tyr 257, Ile 259, Lys 287, Leu 298, Glu 301, Leu 302, Lys 305, particularly Leu 132, Phe 153, Val 157, Val 233, Ile 235, Tyr 257, Ile 259, Lys 287, Leu 298, Glu 301, Leu 302, Lys 305 of LFA-1 I-domain, more particularly by the side chains of such amino acids. In this pocket, the non-hydrogen atoms of mevinolin preferably interact within a distance of  less than 5 xc3x85, particularly 4-4.5 xc3x85.
The complex south pole pocket/mevinolin is energetically favored by hydrophobic, van der Waals and/or electrostatic interactions and possibly also by indirect hydrogen bonding.
As it will be appreciated, there are 2 complexes LFA-1 I-domain/mevinolin per asymemetric unit which are related by a non-crystallographic two fold axis.
As a result of its shape the south pole pocket as defined above favorably associates not only with mevinolin but with other chemical entities or ligands. Such entities or compounds are LFA-1 inhibitors or LFA-1/ICAM-1 or ICAM-3 interaction inhibitors.
The present invention provides any chemical entity or ligand, which binds in whole or in part to the south pole pocket of LFA-1 I-domain as defined above. Preferably the chemical entity or ligand interacts within a distance  less than 5 xc3x85, particularly 4-4.5 xc3x85. Suitable examples of such chemical entities include e.g. mevinolin derivatives. The elucidation of the mevinolin binding interactions on the LFA-1 I-domain south pole pocket provides the necessary information for designing new chemical entities and compounds that may interact in whole or part with the south pole pocket. Thus, the present invention permits the use of molecular design techniques e.g. computer modeling techniques, as a means of identifying, selecting and designing chemical entities or compounds capable of binding to the south pole pocket.
The design of compounds that bind to the south pole pocket according to the invention generally involves consideration of two factors. First, the entity must be capable of physically and structurally associating with parts or all of the south pole pocket. Non-covalent molecular interactions important in this association include hydrophobic, van der Waals interactions, hydrophobic interactions and/or electrostatic interactions and possibly also hydrogen bonding.
Second, the entity must be able to assume a conformation that allows it to associate with the south pole pocket directly. Although certain portions of the entity will not directly participate in these associations, those portions of the entity may still influence the overall conformation of the molecule. This, in turn, may have a significant impact on potency. Such conformational requirements include the overall three-dimensional structure and orientation of the chemical entity in relation to all or a portion of the south pole pocket, or the spacing between functional groups of an entity comprising several chemical entities that directly interact with the south pole pocket.
The chemical entities which interact in whole or in part with the south pole pocket, preferably in a way similar to that of mevinolin may further be tested for their ability to inhibit LFA-1/ICAM-1 or ICAM-3 interactions, using the Jurkat or Hut 78 cell assay described below under A). Representative compounds which bind to the south pole pocket according to the invention are those which inhibit the adhesion of Jurkat or Hut 78 cells to ICAM-1 with an IC50xe2x89xa630 xcexcM. These compounds are indicated as LFA-1 antagonists or LFA-1/ICAM-1 or ICAM-3 interaction inhibitors.
Preferred compounds of the invention for use in accordance with the invention are mevinolins (hereinafter referred to as xe2x80x9cmevinolins of the inventionxe2x80x9d), preferably those having no or only limited HMG CoA R inhibitory activity.
Accordingly, the invention provides:
1. A compound for use in the treatment and/or prevention of autoimmune diseases, acute or chronic inflammatory diseases, ischemia/reperfusion injury, acute or chronic rejection of organ or tissue allo- or xenografts or infection diseases by virtue of its LFA-1 inhibitory activity.
1.1 A compound for use in the treatment and/or prevention of autoimmune diseases, acute or chronic inflammatory diseases, ischemia/reperfusion injury, acute or chronic rejection of organ or tissue allo- or xenografts or infection diseases, the compound binding in whole or part to the south pole pocket, e.g. as defined above, e.g. with an interaction at a distance  less than 5 xc3x85, preferably 4-4.5 xc3x85.
1.2 Mevinolins for use in the treatment and/or prevention of autoimmune diseases, acute or chronic inflammatory diseases, ischemia/reperfusion injury, acute or chronic rejection of organ or tissue allo- or xenografts or infection diseases, by virtue of their LFA-1 inhibitory activity.
1.3 Mevinolins for use in the treatment and/or prevention of autoimmune diseases, acute or chronic inflammatory diseases, ischemia/reperfusion injury, acute or chronic rejection of organ or tissue allo- or xenografts or infection diseases, which bind in whole or in part to the south pole pocket, e.g. as defined above, e.g. with an interaction at a distance  less than 5 xc3x85, preferably 4-4.5 xc3x85.
1.4 Mevinolins for use in the treatment and/or prevention of autoimmune diseases, acute or chronic inflammatory diseases, ischemia/reperfusion injury, acute or chronic rejection of organ or tissue allo- or xenografts or infection diseases, according to 1.2 or 1.3, which inhibit HMG CoA R activity with an IC50xe2x89xa71 xcexcM in the In Vitro Microsomal Assay of HMG CoA R Inhibition as disclosed below.
2. A method for producing a chemical entity or ligand which associates with the LFA-1 I-domain south pole pocket comprising the steps of:
a. employing computational means to perform a fitting operation between the chemical entity and the south pole pocket; and
b. analyzing the results of said fitting operation to quantify the association between the chemical entity and the south pole pocket.
200 xcexcl aliquots (1.08-1.50 mg/ml) of rat liver microsomal suspensions, freshly prepared from male Spargue-Dawley rats (150-225 g body weight), in Buffer A with 10 mmol dithiothreitol are incubated with 10 xcexcl test substance dissolved in dimethylacetamide and assayed for HMG CoA R activity as described by Ackerman et al., J Lipid Res. 18, 408-413 (1977). In the assay the microsomes are the source of the HMG CoA R enzyme which catalyses the reduction of HMG CoA R to mevalonate. The assay employs a chloroform extraction to separate the product, [14C]mevalonolactone, formed by the HGM CoA R reaction from the substrate, [14C]HMG-CoA. [3H]mevalono-lactone is added as an internal reference. Inhibition of HMG CoA R is calculated from the decrease in specific activity [14C/3H]mevalonate of test substances compared to controls and is expressed as IC50 (concentration of test substance which inhibits 50% of HMG CoA R activity).
The utility of the compounds of the invention, e.g. the mevinolins of the invention as inhibitors of LFA-1/ICAM-1 or ICAM-3 interactions may be demonstrated in following test methods:
A. In Vitro
Jurkat or Hut 78 cells obtained from ATCC and cultured in RPMI-1640 supplemented with 10% FCS, L-glutamine, non essential amino acids and 0.05 mM 2-mercaptoethanol, are centrifuged, washed once in PBS, and resuspended at 0.5xc3x97106 cells/ml in binding buffer (1.5% BSA, 5 mM glucose, 2 mM MgCl2, 2ml MnCl2 in TBS, pH 7) containing 5 xcexcg/ml BCECF-AM (Molecular Probes). The cells are incubated at 37xc2x0 C. for 30-45 min. in the dark. Then the cells are centrifuged and resuspended in binding buffer by pipetting and immediately used for experiment.
Flat well microtiter plates (NUNC Maxisorp) are coated with 1 xcexcg/ml goat anti-mouse Cxcexa (Bioreba, South.Biot.) in carbonate buffer (15 mM Na2CO3, 35 mM NaHCO3, pH8.0) 2 hours at 37xc2x0 C. The plates are emptied and blocked with 1.5% BSA and 0.5% Tween-20 in carbonate buffer for 90 min. at 37xc2x0 C. The plates are emptied and washed once in TBS containing, 1.5% BSA. Baculovirus derived ICAM-1 mouse Cxcexa fusion protein (100 ng/ml in TBS/1.5% BSA) is added to the wells. The plates are incubated for 90 min. at 37xc2x0 C. After three washes with TBS/1.5% BSA, the compound to be tested is diluted in binding buffer (as above but free from BSA and glucose) and added to the wells. Then 100 000 Jurkat or Hut 78 cells/well are added and allowed to adhere for 30 min. at 37xc2x0 C. Adherent cells are separated from non-adherent cells by 2-4 washes using binding buffer. Adherent cells are quantified with a fluorescence ELISA reader CytofluorII with the filters set at 485 nm and 530 nm emission.
In this assay, compounds of the invention inhibit adhesion of the Jurkat or Hut 78 cells to ICAM-1 with an IC50xe2x89xa630 xcexcM, preferably 0.05 to 30 xcexcM.
B. In Vivo
i) Murine Thioglycollate Induced Peritonitis
Thioglycollate is injected i.p. to mice and immediately thereafter the compound to be tested is given s.c. The mice are killed after 4 hours, the peritoneal cavity lavaged and the total number of neutrophils in the lavage fluid is determined.
In this assay, the compounds of the invention, e.g. mevinolins inhibit thioglycollate induced neutrophil migration when administered s.c. at a dose of from 0.001-50 mg/kg.
ii) Ischemia/Reperfusion Injury
The compounds may be tested in a model of heart ischemia/reperfusion injury (Abdeslam Oubenaissa et al., Circulation, 94, Suppl. II, 254-258, 1996) or as follows:
Mice weighing 20-25 g are anaesthetized with isoflurane and the right renal vessels are clamped using microvascular clamps for 60 min. After 60 min of ischemia, the microvascular clamps are removed. The left renal vessels (renal artery, vein and ureter) are ligated using a 4-0surgical suture. The left (nonischemic) kidney is removed, and the abdominal cavity closed with 3-0 surgical suture. Sham groups undergo the same procedures as the ischemia group, but without clamping of the reight renal vessels.
Animals are sacrified by CO2 inhalation at 24 h, 1 week and 2 weeks following reperfusion. Blood samples are collected by cardiac puncture into a 3.0 ml Vacutainer(copyright) tube (Becton-Dickensen) containing 0.04 ml of a 7.5% solutio of K3 EDTA immediately after sacrifice. Plasma is separated and stored at xe2x88x9220xc2x0 C. until further analysis. Plasma creatinine and blood urea nitrogen (BUN) are analysed using Sigma procedures. Following sacrifice, the kidney is flushed with physiological saline, immediately snap-frozen in liquid nitrogen and stored at xe2x88x9270xc2x0 C. until analysis. Myeloperoxidase activity (MPO) in the kydney is measured according to the method of Bradley et al (J. Invest. Dermatol., 78, 206-209, 1982).
In this model, the compounds of the invention, e.g. the mevinolins reduce plasma creatinine and blood urea nitrogen when administered at a dose of 0.001 to 50 mg/kg, particularly for 4 days prior to ischemia.
iii) Vascularized Heterotopic Heart Transplantation
Mice donor hearts are implanted onto the recipients abdominal vessels: brachiocephalic trunk to aorta and right pulmonary artery to inferior vena cava with end-to-side anastomoses using 11/0 Ethilon (Ethicon, Norderstedt, Germany) continuous sutures. Animals are closed in two layers with 6/0 Vicryl (Ethicon) and kept warm until fully recovered. Total ischaemia times are in the range of 40-50 min of which 25-35 min are at 4xc2x0 C. During anastomosis (10-15 min) the graft is kept cold.
After transplantation, graft function is monitored by daily assessment of graft beat (palpation). Rejection is considered to be complete when heart beat stops. In all experiments rejection is confirmed by histological examination of the grafts. Significant improvements of graft function are obtained in animals treated with a compound of the invention, e.g. a mevinolin, administered at daily a dosexe2x89xa650 mg/kg.
The compounds of the invention, e.g. the mevinolins of the invention are, therefore, useful in the treatment and/or prevention of diseases or disorders mediated by LFA-1/ICAM-1 interactions e.g. ischemia/reperfusion injury e.g. myocardial infarction, stroke, gut ischemia, renal failure or hemorrhage shock, acute or chronic rejection of organ or tissue allo- or xenografts, acute or chronic inflammatory or autoimmune diseases, e.g. rheumatoid arthritis, asthma, allergy conditions, dermatological diseases, e.g. psoriasis, contact dermatitis, adult respiratory distress syndrome, inflammatory bowel disease and ophthalmic inflammatory diseases, infection diseases such as septic shock, traumatic shock.
For the above uses the required dosage will of course vary depending on the mode of administration, the particular condition to be treated and the effect desired. In general, however, satisfactory results are achieved at dosage rates of from about 0.5 to 80 mg/kg animal body weight. Suitable daily dosage rates for larger mammals, for example humans, are of the order of from about 20 mg to 1.5 g/day, e.g. 100 mg to 1,5 g/day conveniently administered once, in divided dosages 2 to 4xc3x97/day, or in sustained release form. Unit dosage forms suitably comprise from about 5 mg to 0.750 g of a compound of the invention, together with a pharmaceutical acceptable diluent or carrier therefor.
The mevinolins of the invention may be administered in free form or in pharmaceutically acceptable salt form e.g. acid addition salts or alkali salts such as sodium or potassium, or substituted or unsubstituted ammonium salts. Such salts may be prepared in conventional manner and exhibit the same order of activity as the free compounds.
In accordance with the foregoing the present invention further provides:
3. A method for preventing or treating disorders or diseases mediated by LFA-1/ICAM-1 interactions, e.g. such as indicated above in a subject in need of such treatment, which method comprises administering to said subject an effective amount of a compound of the invention, e.g. a mevinolin of the invention or a pharmaceutically acceptable salt thereof;
4. A pharmaceutical composition for use in the method as in 3) above comprising a compound of the invention, e.g. a mevinolin in free form or pharmaceutically acceptable salt form in association with a pharmaceutically acceptable diluent or carrier therefor.
5. A compound of the invention, e.g. a mevinolin or a pharmaceutically acceptable salt thereof for use in the preparation of a pharmaceutical composition for use in the method as in 3) above.
The pharmaceutical compositions may be manufactured in conventional manner.
The compounds of the invention, e.g. the mevinolins of the invention may be administered by any conventional route, for example enterally, preferably orally, e.g. in the form of tablets or capsules or parenterally e.g. in form of injectable solutions or suspensions, or in a nasal or a suppository form.
The mevinolins of the invention may be administered as the sole active ingredient or together with other drugs in immunomodulating regimens or other anti-inflammatory agents for the treatment or prevention of allo- or xenograft acute or chronic rejection or inflammatory or autoimmune disorders. For example, they may be used in combination with cyclosporins, rapamycins or ascomycins, or their immunosuppressive analogs, e.g. cyclosporin A, cyclosporin G, FK-506, rapamycin, 40-O-(2-hydroxy)ethyl-rapamycin etc.; corticosteroids; cyclophosphamide; azathioprene; methotrexate; brequinar; FTY 720; leflunomide; mnizoribine; mycophenolic acid; mycophenolate mofetil; 15-deoxyspergualine; immunosuppressive monoclonal antibodies, e.g., monoclonal antibodies to leukocyte receptors, e.g., MHC, CD2, CD3, CD4, CD7, CD25, CD28, B7, CD45, or CD58 or their ligands; or other immunomodulatory compounds, e.g. CTLA4Ig, or other adhesion molecule inhibitors, e.g. mAbs or low molecular weight inhibitors including Selectin antagonists and VLA-4 antagonists.
Where the mevinolins of the invention are administered in conjunction with other immunosuppressive/immunomodulatory or anti-inflammatory therapy, e.g. for preventing or treating chronic rejection as hereinabove specified, dosages of the co-administered immunosuppressant, immunomodulatory or anti-inflammatory compound will of course vary depending on the type of co-drug employed, e.g. whether it is a steroid or a cyclosporin, on the specific drug employed, on the condition being treated and so forth. In accordance with the foregoing the present invention provides in a yet further aspect:
6. A method as defined above comprising co-administration, e.g. concomitantly or in sequence, of a therapeutically effective amount of a mevinolin of the invention in free form or in pharmaceutically acceptable salt form, and a second drug substance, said second drug substance being an immunosuppressant, immunomodulatory or anti-inflammatory drug, e.g. as indicated above.
7. A kit or package for use in any method as defined under 3) above, comprising a mevinolin of the invention, in free form or in pharmaceutically acceptable salt form, with at least one pharmaceutical composition comprising an immunosuppressant, immunomodulatory or anti-inflammatory drug. The kit or package may comprise instructions for its administration.
Representative mevinolins for use in accordance with the invention are those comprising a moiety of formula A 
which is further substituted in positions 4 and 5 and optionally in positions 6 or 7, each of a-b and xcex1-xcex2, independently being a single or double bond. Each of a-b and xcex1-xcex2 may also be part of a cyclopropyl group. The moiety of formula A may be substituted in positions 4, 5, 6 and/or 7 with one or more substituents as described in literature for known mevinolins, e.g. as disclosed by Y. Chapleur (see above). Preferably the substituent in position 4 is a substituted methyl group. Preferably the substituent in position 5 is linked via xe2x80x94*Oxe2x80x94COxe2x80x94 to the bicyclic residue; more preferably it is xe2x80x94*Oxe2x80x94COxe2x80x94R2 wherein
R2 is C1-C8 alkyl, C3-7 cycloalkyl, aryl, heteroaryl, C3-7 cycloalkyl-C1-4 alkyl, aryl-C1-4 alkyl or heteroaryl-C1-4 alkyl.
Preferred mevinolins of the invention for use as LFA-1 antagonist are compounds of formula I 
wherein
R2 is as defined above, 
Ra is H; C1-6 alkyl; C1-6 alkyl substituted by OH or C1-4 alkoxy; C2-6 alkenyl; or aryl-C1-4 alkyl;
R3 is a radical of formula (i), (ii), (iii) or (iv) 
xe2x80x83wherein X1 and Y1 are (H,H), (H,OH) or xe2x95x90O;
X2 and Y2 are xe2x95x90O or (R,R) wherein each R independently is H, C1-3 alkyl, substituted C1-3 alkyl or X2 and Y2 form together with the cabon atom to which they are bound a 4-, 5-, 6- or 7- membered carbo- or heterocyclic residue,
R4 is ORa wherein Ra is as defined above; or xe2x80x94Oxe2x80x94CORb wherein Rb is C1-8 alkyl optionally substituted by OH, C3-7 cycloalkyl, C3-7 cycloalkyl-C1-4 alkyl, aryl, aryl-C1-4 alkyl, heteroaryl or heteroaryl-C1-4 alkyl; or NRcRd wherein each of Rc and Rd, independently, is C1-6alkyl or form together with the nitrogen to which they are bound a heterocyclic radical optionally comprising an oxygen or another nitrogen atom;
R5 is H, C1-4 alkyl, C3-9 alkenyl, C3-9 alkynyl, aryl-C1-4 alkyl, or C3-7 cycloalkyl-C1-4 alkyl;
R6 is xe2x80x94CHR11xe2x80x94COxe2x80x94NR12R13 wherein R11 has one of the significances as given for R5 and each R12 and R13, independently, is H, C1-4 alkyl, or substituted C1-4 alkyl;
R7 is xe2x95x90O or (H,OH);
R8 is ORa; or NReRf wherein each of Re and Rf, independently, is H, C1-6alkyl, C1-6alkyl substituted by OH or C1-4alkoxy, or a 5-membered heterocyclic residue;
or R7 and R8 together form a dioxy-C1-4alkylene group or xe2x80x94Oxe2x80x94COxe2x80x94Oxe2x80x94;
R9 has one of the significances given for R5;
R10 is COORa; CH2ORc wherein Rc is Ra or CORb; or CONR14R15 or CH2NR14R15 wherein each of R14 and R15 independently is C1-4alkyl, hydroxy substituted C1-4alkyl, carbamoyl-methyl, (C1-4alkyl)-carbamoyl-methyl or di(C1-4alkyl)-carbamoyl-methyl, or one of R14 and R15 is hydrogen and the other is C1-6alkyl, C1-6alkyl substituted by OH and/or a group selected from carbamoyl, (C1-4alkyl)-carbamoyl, di(C1-4alkyl)-carbamoyl and heteroaryl-C1-4alkyl, C1-6alkoxy-carbonyl-methyl, adamantyl-methyl, C3-7cycloalkyl-C1-4alkyl, aryl-C1-4alkyl wherein aryl may be substituted and C1-4alkyl may be substituted by carbamoyl or C1-4alkoxy-carbonyl, or heteroaryl-C1-4alkyl wherein heteroaryl may be substituted by carbamoyl or C1-4alkoxy-carbonyl and C1-4alkyl may be substituted by carbamoyl, or R14 and R15 form together with the nitrogen to which they are attached a heterocyclic residue optionally comprising a further nitrogen atom and optionally substituted by C1-4alkyl, (C1-4alkoxy)-carbonyl, carbamoyl, dioxy-C1-4alkylene, aryl-C1-4alkyl or heteroaryl wherein heteroaryl may be substituted by C1-4alkoxy-carbonyl;
R16 is H; C1-4alkyl; aryl-C1-4alkyl wherein aryl may be substituted by halogen, OH, amino optionally substituted, COOH, CF3, C1-4 alkoxy or cyano; or C3-7cycloalkyl-C1-4aryl;
each of a-b and xcex1-xcex2 independently, is either a single bond or a double bond, in free form or in a pharmaceutically acceptable salt form.
Alkyl groups as Ra Rb, R2, R5, R11, R12 or R13 or alkyl moieties may be branched or straight chain. When R, R12 or R13 is a substituted alkyl, the substituent is preferably located at the end of the alkyl chain and may be e.g. halogen, OH, C3-7 cycloalkyl or aryl. When Re or Rf is substituted C1-6alkyl, it is preferably substituted at the end of the alkyl chain.
Cycloalkyl groups or moieties are preferably cyclopentyl or cyclohexyl.
Aryl or aryl moiety is preferably phenyl and may be substituted, e.g. by halogen, OH, amino optionally substituted, COOH, CF3, C1-4 alkoxy or cyano, preferably by 1,2 or 3 C1-4alkoxy. Aryl-C1-4 alkyl is preferably phenyl-C1-4 alkyl, e.g. benzyl or phenethyl.
Heteroaryl is preferably derived from a 5- or 6-membered heterocycle optionally fused to a benzene ring, e.g. pyrrolyl, imidazolyl, furyl, thienyl, pyridyl, indolyl etc. When Rc and Rd form together with the nitrogen to which they are attached a heterocyclic radical, this may be a 5- or 6-membered ring, e.g. pyrrolidinyl, piperidyl, piperazinyl, 4-methyl-piperazinyl. When one of R14 or R15 is heteroaryl-C1-4alkyl, the heteroaryl moiety may be a 5- or 6-membered, optionally fused to a benzene ring or a heterocyclic residue, e.g. furyl, morpholino, piperazinyl or indolyl. When R14 and R15 form together with the nitrogen to which they are attached a heterocyclic residue, this may be e.g. pyrrolidinyl, piperidino, piperazinyl.
When X2 and Y2 form a carbo- or heterocyclic residue, it may be e.g. cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclohexenyl, pyrrolidinyl, pyrrolidonyl.
The alkylene moiety in dioxy-C1-4alkylene may be linear, e.g. xe2x80x94CH2xe2x80x94, xe2x80x94CH2xe2x80x94CH2xe2x80x94, or branched, e.g. xe2x95x90C(CH3)2.
Compounds of formula I may exist in free form or in salt form, e.g. acid addition salts with e.g. organic or inorganic acids, for example, hydrochlorides, or salt forms obtainable when a COOH is present, as salts with bases e.g. alkali salts such as sodium or potassium, or substituted or unsubstituted ammonium salts.
It will be appreciated that in the residues of formulae (i), (ii) and (iii) the carbon atoms bearing R4, R5, R6, R7, and R8 may be asymetric. Where in the molecule of formula I the stereochemistry is not indicated, it is to be understood that the present invention embraces all enantiomers and their mixtures. Similar considerations apply in relation to starting materials exhibiting asymetric carbon atoms as mentioned above.
In the compounds of formula I, the following significances are preferred:
1. R1 is H or CH3, preferably CH3;
2. R2 is C4-8alkyl, preferably xe2x80x94CH(CH3)xe2x80x94CH2xe2x80x94CH3; xe2x80x94CH(CH2xe2x80x94CH2xe2x80x94CH3)2xe2x80x94; xe2x80x94CH(CH2CH3)2; 
3. R3 is a radical of formula (i);
4. R3 is a radical of formula (iii);
5. R3 is a radical of formula (iii) wherein R7 is (H,OH);
6. R3 is a radical of formula (iii) wherein R7 is xe2x95x90O;
7. R3 is a radical of formula (iii) wherein R8 is OH;
8. R3 is a radical of formula (iii) wherein R7 and R8 form together a dioxy-C1-4alkylene group or xe2x80x94Oxe2x80x94COxe2x80x94Oxe2x80x94;
9. R3 is a radical of formula (iii) wherein R8 is NReRf;
10. R3 is a radical of formula (iii) wherein R8 is NHRf wherein Rf is C1-6alkyl optionally substituted by OH or C1-4alkoxy;
11. R3 is a radical of formula (iii) wherein R9 is H, CH3, benzyl or propargyl;
12. R3 is a radical of formula (iii) wherein R10 is CONR14R15;
13. R3 is a radical of formula (iii) wherein R10 is CONHR15 wherein R15 is C1-4alkyl optionally substituted by OH;
14. R3 is a radical of formula (iii) wherein R10 is CONHR15 wherein R15 is phenyl-C1-4alkyl or heteroaryl-C1-4alkyl wherein the phenyl, heteroaryl and C1-4alkyl moieties may be substituted as indicated above. Preferably phenyl is substituted by 1, 2 or 3 C1-4alkoxy, particularly OCH3;
15. R15 is CH2-phenyl or CH(COxe2x80x94OCH3)-phenyl wherein phenyl may be substituted by 1,2 or 3 C1-4alkoxy, preferably OCH3;
16. R15 is CH2-furyl or CH(CONH2)xe2x80x94CH2-3-indolyl;
17. R10 is xe2x80x94CONR14R15 wherein R14 and R15 form together with the nitrogen to which they are attached an optionally substituted piperidinyl group, e.g. substituted by dioxy-C1-4alkylene, preferably dioxy-ethylene;
18. R3 is a radical of formula (iv).
Among the mevinolins of formula I, the compounds of formula II 
wherein
R1, R2, R3 and the doted lines a-b and xcex1-xcex2 are as defined above,
provided that
1) R2 is other than C1-5 alkyl or aryl-C1-4 alkyl when R1 is H, CH3 or C2H5 and R3 is a radical of formula (i) wherein R4 is OH or OCH3, R5 is H or C1-4 alkyl, and X1 and Y1 are xe2x95x90O, or
2) R2 is other than C1-5 alkyl when R3 is a radical of formula (iii) wherein R9 is H and R10 is COORa,
or salt thereof, are novel and form part of the present invention.
Particularly preferred compounds of formula II are those wherein
1. R1 is H or CH3, preferably CH3 
2. R2 is C4-8alkyl, preferably as disclosed above;
3. R3 is a radical of formula (iii);
4. R3 is a radical of formula (iii) wherein R8 is NHRf wherein Rf is C1-6alkyl optionally substituted by OH or C1-4alkoxy;
5. R3 is a radical of formula (ii) wherein R10 is CONHR15 wherein R15 is C1-4alkyl optionally substituted by OH, preferably OH substituted C1-4alkyl;
6. R3 is a radical of formula (iii) wherein R10 is CONHR15 wherein R15 is phenyl-C1-4alkyl wherein the phenyl moiety may be substituted by 1, 2 or 3 C1-4alkoxy, preferably OCH3.
The present invention also includes a process for the production of the compounds of formula II, comprising
a) for the production of a compound of formula II wherein R3 is a radical of formula (i) or (ii) reacting a compound of formula III 
wherein a-b and xcex1-xcex2 are as defined above, Rxe2x80x21, has one of the significances given for R1 except that the OH group as R1 has to be in protected form and Rxe2x80x23 is a radical of formula (i) or (ii)
with a compound of formula IV
R2 COOHxe2x80x83xe2x80x83IV
wherein R2 is as defined above, or a functional derivative thereof, or
b) converting mevinolin or compactin into a compound of formula I;
and, where required, removing the protecting group, and recovering the compounds of formula II thus obtained in free form or in salt form.
Where OH groups are present in the starting products which are not to participate in the reaction, they may be protected, in accordance with known methods. OH protecting groups are known in the art, e.g. t.-butyl-dimethyl-silanyl.
Process step a) may be performed in accordance with known esterification methods. A functional derivative of a compound of formula IV includes e.g. an acid halogenide, ester or anhydride.
Process step b) may be a substitution in position 5 or a reduction of the pyranyl residue, e.g. as disclosed in Example 3. The R2xe2x80x94COxe2x80x94Oxe2x80x94 group of mevinolin may also be reduced to OH and then esterified to another R2xe2x80x94COxe2x80x94Oxe2x80x94 group. To produce compounds of formula II wherein R3 is a residue of formula (iii) where R10 is CONR14R15, a compound of formula II wherein R3 is a radical of formula (i) or (ii) e.g. mevinolin or compactin, may be submitted to ring opening, e.g. by reaction with a corresponding amine, e.g. alkylamine, HO-alkyl-amine, heterocyclic amine, or via the azide route. When R7 is (H,OH), it may be oxidized to xe2x95x90O in accordance with known oxidation methods, e.g. with sulfur trioxide in form of a pyridine complex or according to a Swern oxidation. The preparation of compounds of formula II wherein R7 and R8 in the radical of formula (iii) form together a carbonate or dioxy-alkylene group, may be performed according to known methods, e.g. using preferably carbonyldiimidazole for the carbonate, or via ketal formation for the dioxy-alkylene group.
Process step b) may also be a cyclisation of a compound of formula II wherein R3 is a radical of formula (iii) to produce a compound of formula II wherein R3 is a radical of formula (i) or (iv). The cyclisation may advantageously be performed in the presence of a base e.g. Hunig""s base and an activating agent, e.g. trifluoromethane sulfonic anhydride. The preparation of a compound of formula II wherein R3 is a radical of formula (iv) may conveniently be performed using a compound of formula II wherein R3 is a radical of formula (iii) wherein R10 is CONHR15 and R7 is oxidized to xe2x95x90O. Cyclisation may be carried out by acidic treatment, e.g. using trifluoroacetic acid.
Insofar as the production of the starting materials is not particularly described, the compounds are known or may be prepared analogously to methods known in the art, as disclosed by Y. Chapleur, in xe2x80x9cProgress in the Chemical Synthesis of Antibiotics and Related Microbial Productsxe2x80x9d, Springer Verlag, 1993, vol. 2, 829-93.
The present invention further provides:
8. A compound of formula II or a pharmaceutically acceptable salt thereof for use as a pharmaceutical, e.g. in the treatment or prevention of disorders or diseases as indicated above.
9. A pharmaceutical composition comprising a compound of formula II, or a pharmaceutically acceptable salt thereof in association with a pharmaceutically acceptable diluent or carrier therefor.